Isolation Technology of Single Spore and Optimization of Conidia Culture Condition for Ustilaginoidea virens

The single spores were isolated from chlamydospores of Ustilaginoidea virens with three different maturities by PSA. The isolated single spores were cul- tured on different media at different temperatures under natural light for inducing conidia to explore the optimum isolation technique of single spore and optimum cul- ture condition of conidia. The results showed that the successful isolating rate of single spore from yellow rice false smut balls reached 90.00%. The sporulatina quantities of isolated single spores cultured on PSD and PDB media at 22 -29 ~C （variable temperature under natural light） or 28 ℃ （constant temperature under dark condition） for 12 d were up to 6.3× 107 and 1.1× 106 spore/mL, respectively. PSA was the most effective method to isolate single spores from yellow rice false smut balls of U. virens. The optimum conidia culture condition included PSD or PDB medium, 22 -29℃ or 28℃, natural light and vibration culture. Key words Ustilaginoidea virens; Single spore isolation; Conidia; Culture condition

Whole lifecycle observation of single‐spore germinated Streptomyces using a nanogap‐stabilized microfluidic chip

Streptomyces is a model bacterium to study multicellular differentiation and the major reservoir for antibiotics discovery.However,the cellular‐level lifecycle of Streptomyces has not been well studied due to its complexity and lack of research tools that can mimic their natural conditions.In this study,we developed a simple microfluidic chip for the cultivation and observation of the entire lifecycle of Streptomyces development from the single‐cell perspective.The chip consists of channels for loading samples and supplying nutrients,microwell arrays for the seeding and growth of single spores,and air chambers beside the microwells that facilitate the development of aerial hyphae and spores.A unique feature of this chip is that each microwell is surrounded by a 1.5µm nanogap connected to an air chamber,which provides a stabilized water–air interface.We used this chip to observe the lifecycle development of Streptomyces coelicolor and Streptomyces griseus germinated from single spores,which revealed differentiation of aerial hyphae with progeny spores at micron‐scale water–air interfaces and air chambers.Finally,we demonstrated the applicability of this chip in phenotypic assays by showing that the microbial hormone A‐Factor is involved in the regulatory pathways of aerial hyphae and spore formation.The microfluidic chip could become a robust tool for studying multicellular differentiation,single‐spore heterogeneity,and secondary metabolism of single‐spore germinated Streptomyces.